One approach commonly used for identifying a change that occurs in a cell consists in detecting a target analyte. See for instance international patent application WO 2008140754.
The measurement of analytes that are important players in chemical, biological, physiological, environmental and other systems can lead to more insights into complex mechanisms. Additionally, the real-time detection of those analytes can provide information on their concentration, fate, or even on their physiological function or effect on cells and organisms. A large variety of detection approaches have been developed that are based on spectroscopic methods. In that context, the use of signal-enhancing structures for Raman spectroscopy has raised a large interest, as illustrated by the fractal structures employed by Ginzburg et al. for the detection of antioxidants [see for instance US patent application U.S. 2004023291]. Others focused on the improvement of methods for non-invasive and in-vivo detection of physiological parameters. For instance, Anderson et al. proposed to use the second derivative spectral values of absorption at two wavelengths of irradiation in order to measure the relative concentration of chromophores, such as haemoglobin, in tissue samples [See e.g. European patent applications EP 1 640 710 and EP 0 816 829]. Differently, the need for integrated detection methods with high throughput has led to the development of platforms that combine standard end-point biomolecule assays and “omic” technologies. Such approach has been suggested as a toxicological platform to evaluate the potential toxic effects of nanomaterials on cells [see international patent application WO 2008010843]. On their side, Fang et al. developed a label-free biosensor that relies on the measurement of the directional mass redistribution in adherent layers of cells exposed to a variety of stimuli [International patent application WO 2006108183].
Hydrogen peroxide (H2O2) belongs to the reactive oxygen species (ROS) of great importance in fields such as biology, pharmaceutics, environment, clinical analysis or food manufacturing. Especially in living organisms H2O2 plays an important role as signalling molecule to regulate biological processes but it is also known for its cytotoxic effects. ROS are principally generated by metabolic processes in the mitochondrial electron transport chain and, especially H2O2, is produced as a side product by many enzymatic reactions. Increased production of H2O2 is associated with oxidative stress which is thought to be involved in the development of many incurable deceases including cancer, Parkinson's decease and Schizophrenia, to name a few examples. End-point bioassays based on membrane-permeable fluorescent and chemiluminescent probes are currently the most widely used to evidence oxidative stress and ROS, including H2O2, production in biological cells exposed to different sources of stress. Those optical methods are very sensitive with detections limits in the pM range. However, fluorescent dyes photo bleaching is one of the major drawback of these optical methods. Furthermore, those indirect techniques are invasive due to the requirement of additional chemicals leading to interferences with the system under observation, which is particularly problematic for measurements in biological systems. Recently, fluorescent single-wall carbon nanotube arrays were used to detect at the μM range H2O2 involved in signalling of epidermal growth factor receptors on the membrane of human carcinoma cells. In addition, the spin-trapping electron spin resonance technique enables real-time probing of intra- and extra-cellular free radical ROS via the formation of semi-stable paramagnetic compounds. On the other hand, electrochemical biosensors, with published detection limits on the order of tens of nanomolars, represent a promising alternative offering non-invasive tracking of the ROS released from cell compartments. Those methods allow real-time measurements but the process performance is limited by slow electrode kinetics and large interferences which may occur due to the availability of other electroactive species in real samples.